The present invention relates to a composition and method for measuring the foliar uptake of an agrochemical. More specifically, it pertains to a composition for measuring the foliar uptake of an agrochemical comprising said agrochemical and Congo Red as a tracer; and a method for measuring the foliar uptake of an agrochemical using same.
Hundreds of agrochemicals have been developed and used commercially to protect crops from weeds, pathogens and insects. However, the prolonged use of agrochemicals have induced the appearance of species which are resistant to once effective agrochemicals, and in order to control such resistant weeds, pathogens or insects, ever increasing amounts of agrochemicals have been applied, causing a serious environmental issue as well as a cost problem. Accordingly, besides the need to develop new classes of environmentally friendly agrochemicals, it is required to suppress the application rates of existing agrochemicals by way of improving, for instance, delivery systems through the addition of effective adjuvants thereto.
Examples of adjuvants used to enhance the activity of an agrochemical include a spreader or spreader-sticker which facilitates the adherence of an agrochemical to a subject crop, a rainfasting agent for preventing the loss of an agrochemical due to rainwash, and a penetrant which allows the absorption of a larger than normal amount of an agrochemical by a subject crop.
As a larger amount of an agrochemical is absorbed by plant tissues in a short time, the manifestation of the intended effect of the agrochemical becomes faster and stronger, as in case of a herbicide. In case of a fungicide, its protective and curative effects against a pathogenic disease increase when a suitable penetrant is employed. Further, in case a proper penetrant is added to a pesticide, it is possible to obtain a superior pesticidal effect of killing harmful insects at sites where the pesticide was not directly applied.
For example, B. Terence et al. have reported that dimethomorph, which is used for treating vine plants infected by downy mildew, potato late blight and pepper blight, showed significantly higher curative activity when used in combination with a nonionic surfactant, e.g., lauryl alcohol ethoxylates(Pesticide Science, 46, 199-213 and 355-359(1996)).
Further, it has been reported that the absorption of glyphosate(N-(phosphonomethyl)glycine, a non-selective herbicide) by plants is significantly facilitated when used in combination with Silwet L-77(Hanjung Chemicals, Inc., Korea), an organosilicon surfactant(Megh Singh, et al., Proceedings of Fourth International Symposium on Adjuvants for Agrochemicals, 385-390(1995)).
As described above, environmental and economic issues demand that the application rate of an agrochemical be kept at a minimal level, while maximizing the intended effectiveness thereof in protecting crops. This effectiveness depends on the amount of the agrochemical absorbed by the crops.
A general method for measuring the amount of agrochemicals absorbed by plants is based on the steps of applying a radiolabelled pesticide to a subject plant, washing the plant with a suitable solvent after a fixed time period, and measuring the radioactivity thereof. However, this method has problems of high cost and complexity which requires a special equipment and the attention of a specialist.
Accordingly, there exist no cost-effective methods for quantifying the foliar uptake of an agrochemical.
As to the procedure for screening penetrants which promote the uptake of an agrochemical by a subject crop, there have been reported the following two methods.
The first method comprises treating a subject plant with a mixture of a radiolabelled agrochemical and a penetrant candidate, and tracing the radio label. However, this method has the aforementioned problems of high cost and complexity.
The second method is based on the steps of applying a mixture of an agrochemical and a penetrant candidate to a subject plant and measuring directly the control effect of the agrochemical against weeds, pathogens or harmful insects. However, this method which requires cumbersome long experimental procedures yield only semi-quantitative data.
The present inventors have endeavored to develop a simple and accurate method for measuring the uptake rate of an agrochemical by a plant and a method for selecting a penetrant suitable for a given agrochemical and plant, and have discovered that a novel tracer can be advantageously employed in accomplishing the above objective.
Accordingly, it is an object of the present invention to provide a composition for quantifying the uptake of an agrochemical by a plant.
Another object of the present invention is to provide a method for quantifying the uptake of an agrochemical by a plant by employing said composition.
A further object of the present invention is to provide a method for selecting a penetrant by employing said method.
In accordance with the present invention, there is provided a composition for measuring the foliar uptake of an agrochemical comprising the agrochemical and Congo Red as a tracer which is totally unabsorbable by plants.
Further, the present invention also provides a method for measuring the foliar uptake of an agrochemical comprising the steps of: (a) applying a composition containing said agrochemical and Congo Red to a plant and a control plate, (b) washing the plant and the control plate with a solvent to extract the agrochemical and Congo Red, (c) measuring the concentrations of the agrochemical and Congo Red in the wash extracts, and (d) calculating the foliar uptake of the agrochemical.
The method of the present invention can be applied for the selection of a suitable penetrant which enhances the uptake of a given agrochemical.
The present invention is described in more detail as follows.
An agrochemical or a formulation thereof is dissolved, emulsified or suspended in water or a water-soluble solvent and Congo Red is added thereto as a tracer and mixed thoroughly to obtain the inventive composition for measuring the foliar uptake of the agrochemical. It is preferable to add Congo Red to the agrochemical solution at a lowest possible concentration, provided that it can be quantitatively analyzed by HPLC or spectrophotometry. For instance, Congo Red can be added to the agrochemical solution at a concentration ranging from 1 xcexcg/ml to 1,000 xcexcg/ml. If the concentration of Congo red in the agrochemical solution is higher than that of the agrochemical, penetration of the agrochemical into the plant may be inhibited, thereby biasing the uptake rate.
Exemplary agrochemicals that can be used in the present invention include all herbicides, insecticides, fungicides, plant growth regulators and the like known in the art.
In preparing the agrochemical solution, an available commercial formulation of agrochemical is merely diluted to a desired concentration by a conventional method. In case of a liquid or jelly-like agrochemical, it may be dissolved in a small amount of an organic solvent such as acetone, or dissolved or emulsified in water with the addition of a small amount of a surfactant, if necessary. In case of a solid agrochemical, it may be pulverized and mixed with a surfactant as a dispersing agent.
For the purpose of selecting a penetrant for a specific agrochemical, the inventive composition may further comprise a penetrant candidate material. The penetrant candidate material may be dissolved in water or an organic solvent and added to the inventive composition.
The inventive composition prepared as in section A above is then applied to a plant and to a control plate which does not absorb the agrochemical or the tracer.
Plant species that can be used in the present invention is not limited, but it is preferable to use a plant which is sufficiently matured and showing no sigh of aging.
The use of a control plate serves the purpose of determining the loss of agrochemical due to reasons other than plant uptake. The material of the control plate useful in the present invention is not limited as far as it neither absorbs, decomposes nor inactivates the inventive composition, and a glass plate is a preferred example thereof.
Congo Red is amenable to photodecomposition and, accordingly, all operations inclusive of the spraying of the agrochemical solution, and the storage and washing of the plants and the control plates are preferably carried out under a dim light, while avoiding direct sunlight. Generally, it is preferable to conduct the operations in a room under a fluorescent lighting.
When spraying the inventive composition to plants and control plates, it is preferable to use a spray booth for a uniform deposition of the composition to target surface. It is also preferable to spray a maximum amount of the composition, but not to the extent that the composition flows down the plants. For instance, the spraying amount of the composition may range from 1 to 100 l/ha for broad-leaf plants whose leaves are horizontal and large, e.g., cucumber and pepper, and from 50 to 300 l/ha for monocotyledonous plants whose leaves are erect and small in area, e.g., rice plants and barnyard grass. However, it should be understood that the spraying amount may vary according to the plant species and various other factors.
It is preferable to store the sprayed plants and control plates in a dark place.
A selected number of the plants and control plates sprayed with the inventive composition as in section B are washed with a suitable solvent immediately after the spraying to obtain standard wash extracts that are used in defining base lines for the subsequent measurements. The remainder of the sprayed plants and control plates are kept in a dark place for a predetermined period and then subjected to similar washing to obtain wash extracts containing unabsorbed agrochemical and Congo Red. Suitable solvents for this wash extraction process include water, water-soluble organic solvents such as methanol, ethanol, acetone and acetonitrile, and a mixture thereof, wherein an acetonitrile-aqueous solution is preferred. When an aqueous solution of an organic solvent is used, the ratio of the water-soluble organic solvent to water may be varied from 0:10 (v/v) to 5:5(v/v) depending on the water-solubility of the agrochemical employed. Namely, it is preferable to lower the content of water-soluble organic solvent when the agrochemical has a high water-solubility, while a lower water content is preferable when the agrochemical has a low water-solubility.
In washing the sprayed plants, an exposed part of the plant is cut and put in a test tube containing a suitable washing solution, and the test tube is capped and agitated with bottom-side up at 50 to 80 rpm for 1 to 5 minutes. The sprayed control plates may be put in a petri dish having a suitable size and washed with the same solvent as above.
The concentrations of the agrochemical and Congo Red in the wash extracts obtained in section C are measured by conventional methods, e.g., HPLC, gas chromatography and spectrophotometry.
Subsequently, the foliar uptake of the agrochemical is calculated by the steps of:
(1) calculating the respective concentration ratios of the agrochemical and Congo Red(Conc. of agrochemical/Conc. of Congo Red) for the initial wash extracts of the plant and control plate obtained immediately after the spraying;
(2) calculating the respective concentration ratios of the agrochemical and Congo Red(Conc. of the agrochemical/Conc. of Congo Red) for the wash extracts of the plant and control plate obtained at a predetermined time after the spraying;
(3) dividing the concentration ratio of the agrochemical and Congo Red calculated in (2) by that calculated in (1) for the wash extracts of the plant and control plate, respectively, to obtain the ratio of the agrochemical remaining on the surface of the plant and control plate at the predetermined time after the spraying;
(4) subtracting the ratio of the agrochemical remaining on the surface of the plant from that of the control plate to obtain the fraction the agrochemical absorbed by the plant.
The inventive method for measuring the uptake rate of an agrochemical will be understood more easily with the following exemplary case.
An agrochemical solution comprising 100 ppm of an agrochemical and 100 ppm of Congo Red is prepared and sprayed to two cucumber leaves and two glass plates. Immediately after the spraying, one cucumber leaf and one glass plate are washed with 10 ml of a solvent, respectively, and the concentrations of the agrochemical and Congo Red in the wash extracts are measured by HPLC. On the other hand, the remaining cucumber leaf and glass plate are stored in a dark room for 24 hours and, then, washed with 10 ml of the same solvent. The concentrations of the agrochemical and Congo Red in the wash extracts are also measured by HPLC. The uptake of the agrochemical by cucumber leaves is calculated according to the previously described procedure, as in Table 1.
As discussed above, Congo Red is neither absorbable by plants nor volatilizable. Accordingly, the total amount of the agrochemical lost due to the absorption by a plant and other factors such as evaporation and decomposition can be calculated by comparing the amounts of the agrochemical and Congo Red found on the plant immediately after the spraying and at predetermined time after the spraying of inventive composition. Further, as a glass plate absorbs no material, the ratio of the agrochemical remaining on the glass plate reflects the ratio of the agrochemical lost by reasons other than absorption by the plant. Accordingly, an accurate figure for the foliar uptake of the agrochemical can be easily calculated by comparing the ratios of the agrochemical remaining on the plant and the glass plate.
As previously discussed, the method of the present invention employing Congo Red as a tracer for an agrochemical is useful for accurate assessment of the amount of the agrochemical absorbed by a plant. Further, the inventive method is also useful for the selection of a penetrant suitable for a specific agrochemical and plant. The present invention can be advantageously used in determining the lowest effective application rates of various agrochemicals, thereby creating benefits in terms of both economy and protection of the environment.
The following Examples are intended to further illustrate the present invention without limiting its scope.
Further, percentages given below for solid in solid mixture, liquid in liquid, and solid in liquid are on a wt/wt, vol/vol and wt/vol basis, respectively, unless specifically indicated otherwise.